Selenium-GPX4 axis protects follicular helper T cells from ferroptosis.

Follicular helper T (TFH) cells are a specialized subset of CD4+ T cells that essentially support germinal center responses where high-affinity and long-lived humoral immunity is generated. The regulation of TFH cell survival remains unclear. Here we report that TFH cells show intensified lipid peroxidation and altered mitochondrial morphology, resembling the features of ferroptosis, a form of programmed cell death that is driven by iron-dependent accumulation of lipid peroxidation. Glutathione peroxidase 4 (GPX4) is the major lipid peroxidation scavenger and is necessary for TFH cell survival. The deletion of GPX4 in T cells selectively abrogated TFH cells and germinal center responses in immunized mice. Selenium supplementation enhanced GPX4 expression in T cells, increased TFH cell numbers and promoted antibody responses in immunized mice and young adults after influenza vaccination. Our findings reveal the central role of the selenium-GPX4-ferroptosis axis in regulating TFH homeostasis, which can be targeted to enhance TFH cell function in infection and following vaccination.

Global histone H2B degradation regulates insulin/IGF signaling-mediated nutrient stress.

Eukaryotic organisms adapt to environmental fluctuations by altering their epigenomic landscapes and transcriptional programs. Nucleosomal histones carry vital epigenetic information and regulate gene expression, yet the mechanisms underlying chromatin-bound histone exchange remain elusive. Here, we found that histone H2Bs are globally degraded in Caenorhabditis elegans during starvation. Our genetic screens identified mutations in ubiquitin and ubiquitin-related enzymes that block H2B degradation in starved animals, identifying lysine 31 as the crucial residue for chromatin-bound H2B ubiquitination and elimination. Retention of aberrant nucleosomal H2B increased the association of the FOXO transcription factor DAF-16 with chromatin, generating an ectopic gene expression profile detrimental to animal viability when insulin/IGF signaling was reduced in well-fed animals. Furthermore, we show that the ubiquitin-proteasome system regulates chromosomal histone turnover in human cells. During larval development, C. elegans epidermal cells undergo H2B turnover after fusing with the epithelial syncytium. Thus, histone degradation may be a widespread mechanism governing dynamic changes of the epigenome.

DeepPBI-KG: a deep learning method for the prediction of phage-bacteria interactions based on key genes.

Phages, the natural predators of bacteria, were discovered more than 100 years ago. However, increasing antimicrobial resistance rates have revitalized phage research. Methods that are more time-consuming and efficient than wet-laboratory experiments are needed to help screen phages quickly for therapeutic use. Traditional computational methods usually ignore the fact that phage-bacteria interactions are achieved by key genes and proteins. Methods for intraspecific prediction are rare since almost all existing methods consider only interactions at the species and genus levels. Moreover, most strains in existing databases contain only partial genome information because whole-genome information for species is difficult to obtain. Here, we propose a new approach for interaction prediction by constructing new features from key genes and proteins via the application of K-means sampling to select high-quality negative samples for prediction. Finally, we develop DeepPBI-KG, a corresponding prediction tool based on feature selection and a deep neural network. The results show that the average area under the curve for prediction reached 0.93 for each strain, and the overall AUC and area under the precision-recall curve reached 0.89 and 0.92, respectively, on the independent test set; these values are greater than those of other existing prediction tools. The forward and reverse validation results indicate that key genes and key proteins regulate and influence the interaction, which supports the reliability of the model. In addition, intraspecific prediction experiments based on Klebsiella pneumoniae data demonstrate the potential applicability of DeepPBI-KG for intraspecific prediction. In summary, the feature engineering and interaction prediction approaches proposed in this study can effectively improve the robustness and stability of interaction prediction, can achieve high generalizability, and may provide new directions and insights for rapid phage screening for therapy.

Proteomic and Cellular Characterization of Omicron Breakthrough Infections and a Third Homologous or Heterologous Boosting Vaccination in a Longitudinal Cohort.

The understanding of dynamic plasma proteome features in hybrid immunity and breakthrough infection is limited. A deeper understanding of the immune differences between heterologous and homologous immunization could assist in the future establishment of vaccination strategies. In this study, 40 participants who received a third dose of either a homologous BBIBP-CorV or a heterologous ZF2001 protein subunit vaccine following two doses of inactivated coronavirus disease 2019 vaccines and 12 patients with BA2.2 breakthrough infections were enrolled. Serum samples were collected at days 0, 28, and 180 following the boosting vaccination and breakthrough and then analyzed using neutralizing antibody tests and mass spectrometer-based proteomics. Mass cytometry of peripheral blood mononuclear cell samples was also performed in this cohort. The chemokine signaling pathway and humoral response markers (IgG2 and IgG3) associated with infection were found to be upregulated in breakthrough infections compared to vaccination-induced immunity. Elevated expression of IGKV, IGHV, IL-17 signaling, and the phagocytosis pathway, along with lower expression of FGL2, were correlated with higher antibody levels in the boosting vaccination groups. The MAPK signaling pathway and Fc gamma R-mediated phagocytosis were more enriched in the heterologous immunization groups than in the homologous immunization groups. Breakthrough infections can trigger more intensive inflammatory chemokine responses than vaccination. T-cell and innate immune activation have been shown to be closely related to enhanced antibody levels after vaccination and therefore might be potential targets for vaccine adjuvant design.

Vascular endothelial growth factor receptor 2 as a potential host target for the inhibition of enterovirus replication.

Because host kinases are key regulators of multiple signaling pathways in response to viral infections, we previously screened a kinase inhibitor library using rhabdomyosarcoma cells and human intestinal organoids in parallel to identify potent inhibitors against EV-A71 infection. We found that Rho-associated coiled-coil-containing protein kinase (Rock) inhibitor efficiently suppressed the EV-A71 replication and further revealed Rock1 as a novel EV-A71 host factor. In this study, subsequent analysis found that a variety of vascular endothelial growth factor receptor (VEGFR) inhibitors also had potent antiviral effects. Among the hits, Pazopanib, with a selectivity index as high as 254, which was even higher than that of Pirodavir, a potent broad-spectrum picornavirus inhibitor targeting viral capsid protein VP1, was selected for further analysis. We demonstrated that Pazopanib not only efficiently suppressed the replication of EV-A71 in a dose-dependent manner, but also exhibited broad-spectrum anti-enterovirus activity. Mechanistically, Pazopanib probably induces alterations in host cells, thereby impeding viral genome replication and transcription. Notably, VEGFR2 knockdown and overexpression suppressed and facilitated EV-A71 replication, respectively, indicating that VEGFR2 is a novel host dependency factor for EV-A71 replication. Transcriptome analysis further proved that VEGFR2 potentially plays a crucial role in combating EV-A71 infection through the TSAd-Src-PI3K-Akt pathway. These findings expand the range of potential antiviral candidates of anti-enterovirus therapeutics and suggest that VEGFR2 may be a key host factor involved in EV-A71 replication, making it a potential target for the development of anti-enterovirus therapeutics. IMPORTANCE: As the first clinical case was identified in the United States, EV-A71, a significant neurotropic enterovirus, has been a common cause of hand, foot, and mouth disease (HFMD) in infants and young children. Developing an effective antiviral agent for EV-A71 and other human enteroviruses is crucial, as these viral pathogens consistently cause outbreaks in humans. In this study, we demonstrated that multiple inhibitors against VEGFRs effectively reduced EV-A71 replication, with Pazopanib emerging as the top candidate. Furthermore, Pazopanib also attenuated the replication of other enteroviruses, including CVA10, CVB1, EV-D70, and HRV-A, displaying broad-spectrum anti-enterovirus activity. Given that Pazopanib targets various VEGFRs, we narrowed the focus to VEGFR2 using knockdown and overexpression experiments. Transcriptomic analysis suggests that Pazopanib's potential downstream targets involve the TSAd-Src-PI3K-Akt pathway. Our work may contribute to identifying targets for antiviral inhibitors and advancing treatments for human enterovirus infections.

Acyloxyacyl hydrolase promotes pulmonary defense by preventing alveolar macrophage tolerance.

Although alveolar macrophages (AMs) play important roles in preventing and eliminating pulmonary infections, little is known about their regulation in healthy animals. Since exposure to LPS often renders cells hyporesponsive to subsequent LPS exposures ("tolerant"), we tested the hypothesis that LPS produced in the intestine reaches the lungs and stimulates AMs, rendering them tolerant. We found that resting AMs were more likely to be tolerant in mice lacking acyloxyacyl hydrolase (AOAH), the host lipase that degrades and inactivates LPS; isolated Aoah-/- AMs were less responsive to LPS stimulation and less phagocytic than were Aoah+/+ AMs. Upon innate stimulation in the airways, Aoah-/- mice had reduced epithelium- and macrophage-derived chemokine/cytokine production. Aoah-/- mice also developed greater and more prolonged loss of body weight and higher bacterial burdens after pulmonary challenge with Pseudomonas aeruginosa than did wildtype mice. We also found that bloodborne or intrarectally-administered LPS desensitized ("tolerized") AMs while antimicrobial drug treatment that reduced intestinal commensal Gram-negative bacterial abundance largely restored the innate responsiveness of Aoah-/- AMs. Confirming the role of LPS stimulation, the absence of TLR4 prevented Aoah-/- AM tolerance. We conclude that commensal LPSs may stimulate and desensitize (tolerize) alveolar macrophages in a TLR4-dependent manner and compromise pulmonary immunity. By inactivating LPS in the intestine, AOAH promotes antibacterial host defenses in the lung.

Vaccination Shapes Within-Host SARS-CoV-2 Diversity of Omicron BA.2.2 Breakthrough Infection.

BACKGROUND: Low-frequency intrahost single-nucleotide variants of SARS-CoV-2 have been recognized as predictive indicators of selection. However, the impact of vaccination on the intrahost evolution of SARS-CoV-2 remains uncertain at present. METHODS: We investigated the genetic variation of SARS-CoV-2 in individuals who were unvaccinated, partially vaccinated, or fully vaccinated during Shanghai's Omicron BA.2.2 wave. We substantiated the connection between particular amino acid substitutions and immune-mediated selection through a pseudovirus neutralization assay or by cross-verification with the human leukocyte antigen-associated T-cell epitopes. RESULTS: In contrast to those with immunologic naivety or partial vaccination, participants who were fully vaccinated had intrahost variant spectra characterized by reduced diversity. Nevertheless, the distribution of mutations in the fully vaccinated group was enriched in the spike protein. The distribution of intrahost single-nucleotide variants in individuals who were immunocompetent did not demonstrate notable signs of positive selection, in contrast to the observed adaptation in 2 participants who were immunocompromised who had an extended period of viral shedding. CONCLUSIONS: In SARS-CoV-2 infections, vaccine-induced immunity was associated with decreased diversity of within-host variant spectra, with milder inflammatory pathophysiology. The enrichment of mutations in the spike protein gene indicates selection pressure exerted by vaccination on the evolution of SARS-CoV-2.

Therapeutic vaccination with lentiviral vector in HBV-persistent mice and two inactive HBsAg carriers.

BACKGROUND & AIMS: Immunotherapy for chronic hepatitis B virus (HBV) infection has not yet demonstrated sufficient efficacy. We developed a non-integrative lentiviral-vectored therapeutic vaccine for chronic hepatitis B and tested its antiviral effects in HBV-persistent mice and two inactive HBsAg carriers. METHODS: Lentiviral vectors (LVs) encoding the core, preS1, or large HBsAg (LHBs) proteins of HBV were evaluated for immunogenicity in HBV-naïve mice and therapeutic efficacy in a murine model of chronic HBV infection. In addition, two inactive HBsAg carriers each received two doses of 5×107 transduction units (TU) or 1×108 TU of lentiviral-vectored LHBs (LV-LHBs), respectively. The endpoints were safety, LHBs-specific T-cell responses, and serum HBsAg levels during a 24-week follow-up. RESULTS: In the mouse models, LV-LHBs was the most promising in eliciting robust antigen-specific T cells and in reducing the levels of serum HBsAg and viral load. By the end of the 34-week observation period, six out of ten (60%) HBV-persistent mice vaccinated with LV-LHBs achieved serum HBsAg loss and significant depletion of HBV-positive hepatocytes in the liver. In the two inactive HBsAg carriers, vaccination with LV-LHBs induced a considerable increase in the number of peripheral LHBs-specific T cells in one patient, and a weak but detectable response in the other, accompanied by a sustained reduction of HBsAg (-0.31 log10 IU/ml and -0.46 log10 IU/ml, respectively) from baseline to nadir. CONCLUSIONS: A lentiviral-vectored therapeutic vaccine for chronic HBV infection demonstrated the potential to improve HBV-specific T-cell responses and deplete HBV-positive hepatocytes, leading to a sustained loss or reduction of serum HBsAg. IMPACT AND IMPLICATIONS: Chronic HBV infection is characterized by an extremely low number and profound hypo-responsiveness of HBV-specific T cells. Therapeutic vaccines are designed to improve HBV-specific T-cell responses. We show that immunization with a lentiviral-vectored therapeutic HBV vaccine was able to expand HBV-specific T cells in vivo, leading to reductions of HBV-positive hepatocytes and serum HBsAg.

PegIFN alpha-2a reduces relapse in HBeAg-negative patients after nucleo(s)tide analogue cessation: A randomized-controlled trial.

BACKGROUND & AIMS: Nucleo(s)tide analogue (NUC) cessation can lead to hepatitis B surface antigen (HBsAg) clearance but also a high rate of virological relapse. However, the effect of pegylated interferon alpha-2a (PegIFN-α-2a) on virological relapse after NUC cessation is unknown. Therefore, this study aimed to evaluate the effect of switching from NUC to PegIFN-α-2a treatment for 48 weeks on virological relapse up to week 96. METHODS: In this multicenter randomized-controlled clinical trial, 180 non-cirrhotic patients with HBeAg-negative chronic hepatitis B on continuous NUC therapy for ≥2.5 years, with HBV DNA levels <60 IU/ml, were randomized to discontinue NUC therapy (n = 90) or receive 48 weeks of PegIFN-α-2a treatment (n = 90). Patients were followed up for up to 96 weeks. The primary endpoint was the virological relapse rate up to week 96. RESULTS: Intention-to-treat analysis revealed patients in the interferon monotherapy group had significantly lower cumulative virological relapse rates than the NUC cessation group until week 96 (20.8% vs. 53.6%, p <0.0001). Consistently, a significantly lower proportion of patients in the interferon monotherapy group had virological relapse than those in the NUC cessation group at 48 weeks off treatment (17.8% vs. 36.7%, p = 0.007). The virological relapse rate positively correlated with HBsAg levels in the NUC cessation group. The interferon monotherapy group had a lower cumulative clinical relapse rate (7.8% vs. 20.9%, p = 0.008) and a higher HBsAg loss rate (21.5% vs. 9.0%, p = 0.03) than the NUC cessation group. CONCLUSIONS: Switching from NUC to PegIFN-α-2a treatment for 48 weeks significantly reduces virological relapse rates and leads to higher HBsAg loss rates than NUC treatment cessation alone in patients with HBeAg-negative chronic hepatitis B. IMPACT AND IMPLICATIONS: Nucleo(s)tide analogue (NUC) cessation can lead to HBsAg clearance but also a high rate of virological relapse, but an optimized scheme to reduce the virological relapse rate after NUC withdrawal is yet to be reported. This randomized-controlled trial investigated the effect of switching from NUC to PegIFN-α-2a treatment for 48 weeks on virological relapse up to week 96 in patients with HBeAg-negative chronic hepatitis B. The interferon monotherapy group had a significantly lower cumulative virological relapse rate (20.8% vs. 53.6%, p <0.0001) and higher HBsAg loss rate (21.5% vs. 9.0%, p = 0.03) than the NUC cessation group up to week 96. This provides an optimized strategy for NUC cessation in HBeAg-negative patients. TRIAL REGISTRATION NUMBER: NCT02594293.

The diagnostic value and validation of IL-22 combimed with sCD40L in tuberculosis pleural effusion.

BACKGROUND: There is substantial evidence indicating that cytokines play a role in the immune defense against tuberculosis. This study aims to evaluate the levels of various cytokines in pleural effusion to ditinguish between tuberculosis pleurisy and malignant pleurisy. METHODS: A total of 82 participants with pleural effusion were included in the training cohort, and 76 participants were included in the validation cohort. The individuals were divided into tuberculosis and malignant pleurisy groups. The concentrations of interleukin-1ß (IL-1ß), IL-4, IL-6, IL-10, IL-17 A, IL-17 F, IL-21, IL-22, IL-25, IL-31, IL-33, interferon-γ (IFN-γ), soluble CD40 ligand (sCD40L) and tumor necrosis factor-α (TNF-α) in pleural effusion were measured using a multiplex cytokine assay. The threshold values were calculated according to the receiver operating characteristic (ROC) curve analysis to aid in diagnosing tuberculosis pleurisy. Furthermore, the combined measure was validated in the validation cohort. RESULTS: The levels of all 14 cytokines in pleural effusion were significantly higher in participants with tuberculosis compared to those with malignant pleurisy (all P < 0.05). The area under the curve (AUC) was ≥ 0.920 for the IL-22, sCD40L, IFN-γ, TNF-α and IL-31, which were significantly increased in tuberculous pleural effusion (TPE) compared to MPE in the training cohort. Threshold values of 95.80 pg/mL for IFN-γ, 41.80 pg/mL for IL-31, and 18.87 pg/mL for IL-22 provided ≥ 90% sensitivity and specificity in distinguishing between tuberculosis pleurisy and malignant pleurisy in the training cohort. Among these, IL-22 combined with sCD40L showed the best sensitivity and specificity (94.0% and 96.9%) for diagnosing tuberculosis pleurisy, and this finding was validated in the validation cohort. CONCLUSION: We demonstrated that the levels of IL-1ß, IL-4, IL-6, IL-10, IL-17 A, IL-17 F, IL-21, IL-22, IL-25, IL-31, IL-33, IFN-γ, sCD40L and TNF-α in pleural effusion had significant difference between tuberculosis pleurisy and malignant pleurisy. Specifically, IL-22 ≥ 18.87 pg/mL and sCD40L ≥ 53.08 pg/mL can be clinically utilized as an efficient diagnostic strategy for distinguishing tuberculosis pleurisy from malignant pleurisy.

A Novel Homozygous RHOH Variant Associated with T Cell Dysfunction and Recurrent Opportunistic Infections.

RHOH, an atypical small GTPase predominantly expressed in hematopoietic cells, plays a vital role in immune function. A deficiency in RHOH has been linked to epidermodysplasia verruciformis, lung disease, Burkitt lymphoma and T cell defects. Here, we report a novel germline homozygous RHOH c.245G > A (p.Cys82Tyr) variant in a 21-year-old male suffering from recurrent, invasive, opportunistic infections affecting the lungs, eyes, and brain. His sister also succumbed to a lung infection during early adulthood. The patient exhibited a persistent decrease in CD4+ T, B, and NK cell counts, and hypoimmunoglobulinemia. The patient's T cell showed impaired activation upon in vitro TCR stimulation. In Jurkat T cells transduced with RHOHC82Y, a similar reduction in activation marker CD69 up-regulation was observed. Furthermore, the C82Y variant showed reduced RHOH protein expression and impaired interaction with the TCR signaling molecule ZAP70. Together, these data suggest that the newly identified autosomal-recessive RHOH variant is associated with T cell dysfunction and recurrent opportunistic infections, functioning as a hypomorph by disrupting ZAP70-mediated TCR signaling.

Multi-omics landscapes reveal heterogeneity in long COVID patients characterized with enhanced neutrophil activity.

BACKGROUND: Omicron variant impacts populations with its rapid contagiousness, and part of patients suffered from persistent symptoms termed as long COVID. The molecular and immune mechanisms of this currently dominant global variant leading to long COVID remain unclear, due to long COVID heterogeneity across populations. METHODS: We recruited 66 participants in total, 22 out of 66 were healthy control without COVID-19 infection history, and 22 complaining about long COVID symptoms 6 months after first infection of Omicron, referred as long COVID (LC) Group. The left ones were defined as non-long COVID (NLC) Group. We profiled them via plasma neutralizing antibody titer, SARS-CoV-2 viral load, transcriptomic and proteomics screening, and machine learning. RESULTS: No serum residual SARS-CoV-2 was observed in the participants 6 months post COVID-19 infection. No significant difference in neutralizing antibody titers was found between the long COVID (LC) Group and the non-long COVID (NLC) Group. Transcriptomic and proteomic profiling allow the stratification of long COVID into neutrophil function upregulated (NU-LC) and downregulated types (ND-LC). The NU-LC, identifiable through a refined set of 5 blood gene markers (ABCA13, CEACAM6, CRISP3, CTSG and BPI), displays evidence of relatively higher neutrophil counts and function of degranulation than the ND-LC at 6 months after infection, while recovered at 12 months post COVID-19. CONCLUSION: The transcriptomic and proteomic profiling revealed heterogeneity among long COVID patients. We discovered a subgroup of long COVID population characterized by neutrophil activation, which might associate with the development of psychiatric symptoms and indicate a higher inflammatory state. Meanwhile, a cluster of 5 genes was manually curated as the most potent discriminators of NU-LC from long COVID population. This study can serve as a foundational exploration of the heterogeneity in the pathogenesis of long COVID and assist in therapeutic targeting and detailed epidemiological investigation of long COVID.

A single center retrospective study: Comparison between centrifugal separation plasma exchange with ACD-A and membrane separation plasma exchange with heparin on acute liver failure and acute on chronic liver failure.

The purpose of this retrospective study is to compare the efficacy and safety of the centrifugal separation therapeutic plasma exchange (TPE) using citrate anticoagulant (cTPEc) with membrane separation TPE using heparin anticoagulant (mTPEh) in liver failure patients. The patients treated by cTPEc were defined as cTPEc group and those treated by mTPEh were defined as mTPEh group, respectively. Clinical characteristics were compared between the two groups. Survival analyses of two groups and subgroups classified by the model for end-stage liver disease (MELD) score were performed by Kaplan-Meier method and were compared by the log-rank test. In this study, there were 51 patients in cTPEc group and 18 patients in mTPEh group, respectively. The overall 28-day survival rate was 76% (39/51) in cTPEc group and 61% (11/18) in mTPEh group (P > .05). The 90-day survival rate was 69% (35/51) in cTPEc group and 50% (9/18) in mTPEh group (P > .05). MELD score = 30 was the best cut-off value to predict the prognosis of patients with liver failure treated with TPE, in mTPEh group as well as cTPEc group. The median of total calcium/ionized calcium ratio (2.84, range from 2.20 to 3.71) after cTPEc was significantly higher than the ratio (1.97, range from 1.73 to 3.19) before cTPEc (P < .001). However, there was no significant difference between the mean concentrations of total calcium before cTPEc and at 48 h after cTPEc. Our study concludes that there was no statistically significant difference in survival rate and complications between cTPEc and mTPEh groups. The liver failure patients tolerated cTPEc treatment via peripheral vascular access with the prognosis similar to mTPEh. The prognosis in patients with MELD score < 30 was better than in patients with MELD score ≥ 30 in both groups. In this study, the patients with acute liver failure (ALF) and acute on chronic liver failure (ACLF) treated with cTPEc tolerated the TPE frequency of every other day without significant clinical adverse event of hypocalcemia with similar outcomes to the mTPEh treatment. For liver failure patients treated with cTPEc, close clinical observation and monitoring ionized calcium are necessary to ensure the patients' safety.

Influence of Dyadic Coping Model on Anxiety and Depression Level and Treatment Compliance in Glaucoma Patients.

Objective: To investigate the influence of the dyadic coping model on anxiety and depression levels and treatment compliance in glaucoma patients. Methods: According to the random number table method, 80 glaucoma patients were assigned into an observation group and a control group, with 40 cases in each group are recruited from January 2021 to February 2022. Both groups received routine preoperative glaucoma care; in addition, the observation group received a 10-week dyadic coping model intervention. The dyadic coping model is a therapeutic approach that involves the collaborative efforts of both patients and their close partners or caregivers to cope with stressors and challenges related to the perioperative period. The baseline data questionnaires were collected before the intervention, and the outcome was evaluated 10 weeks later using the Anxiety and Depression Self-Rating Scale and the Treatment Compliance Scale. Results: After intervention, the treatment compliance of glaucoma in the observation group was significantly better than that in the control group, and the anxiety and depression level in the observation group was significantly lower than that in the control group (P < .05). Conclusion: The dyadic coping model intervention for glaucoma patients can successfully increase treatment compliance and lower anxiety and depression levels.

Clinical Characteristics, Treatment, and Prognosis of Osteoarticular Brucellosis: A Retrospective Real-World Study in Shenyang, China, 2014-2019.

This study aimed to assess the clinical characteristics, treatment, and prognosis of osteoarticular brucellosis. We conducted a retrospective study enrolling brucellosis patients from the Sixth People's Hospital of Shenyang between September 2014 and June 2019. A total of 1917 participants were admitted during this period. After applying propensity score matching, we retrospectively analyzed 429 patients with osteoarthritis and 429 patients without osteoarthritis. The primary outcome was treatment completion. The secondary outcome was symptom disappearance and seroconversion. Brucellosis patients with osteoarthritis had longer treatment course (160 [134.3-185.7] vs. 120 [102.3-137.7] d, p = 0.008) than those without osteoarthritis. The most common involved site was lumbar vertebrae (290 [67.6%]) in brucellosis patients with osteoarthritis. Longer symptom duration (90 [83.0-97.0] vs. 42 [40.2-43.8], p < 0.001) along with no significant difference in seroconversion (180 [178.8-181.2] vs. 180 [135.1-224.9], p = 0.212) was observed in osteoarthritis patients with treatment course >90 d. Peripheral joint involvement (adjusted hazard ratio [95% confidence interval] 1.485 [1.103-1.999]; p = 0.009) had a shorter symptom duration compared with shaft joint involvement. No significant differences were observed in treatment therapy between doxycycline plus rifampin (DR) or plus cephalosporins (DRC) in treatment course (p = 0.190), symptom persistence (p = 0.294), and seroconversion (p = 0.086). Lumbar vertebra was the most commonly involved site. Even if all symptoms disappeared, Serum agglutination test potentially remained positive in some patients. Compared with peripheral arthritis, shaft arthritis was the high-risk factor for longer symptom duration. The therapeutic effects were similar between DR and DRC. In summary, our study provided important insights into the clinical characteristics, treatment, and outcomes of osteoarticular brucellosis. Clinical Trial Registration number: NCT04020536.

Evidence of Residual Ongoing Viral Replication in Chronic Hepatitis B Patients Successfully Treated With Nucleos(t)ide Analogues.

BACKGROUND: Chronic hepatitis B is usually treated with nucleos(t)ide analogues (NAs). However, a cure is rarely achieved, even with years of treatment. Here, we investigated whether viral replication is completely halted and how long covalently closed circular DNA (cccDNA) persists in patients successfully treated with NAs. METHODS: A series of longitudinal serum samples and a collection of cross-sectional liver biopsies were obtained from patients successfully treated with NAs. Viral variants in serum HBV RNA were enumerated by deep sequencing. Viral replication intermediates in hepatocytes were directly visualized by in situ hybridization. The apparent half-life of each cccDNA was estimated. RESULTS: Three of 6 successfully treated patients demonstrated clear evidence of a small proportion of virus evolution, although the overwhelming proportion of variants were identical or possessed a similar degree of divergence through time. The apparent half-life of variants was estimated to be from approximately 7.42 weeks to infinite. Hepatocytes remained positive for cytoplasmic nucleocapsids-associated relaxed circular DNA in 4 of 7 liver needle biopsies. CONCLUSIONS: We conclude that even after prolonged treatment, a small proportion of the cccDNA reservoir is constantly replenished by continued low-level HBV replication, whereas a large proportion of the cccDNA reservoir persists over time.

Single-Ascending-Dose, Food-Effect, and Multiple-Dose Study of the Safety, Tolerability, and Pharmacokinetics of the Pangenotypic Anti-Hepatitis C Virus Drug Holybuvir in Healthy Chinese Subjects.

Holybuvir is a novel pangenotypic hepatitis C virus NS5B inhibitor. This first in-human study aimed to evaluate the pharmacokinetics (PK), safety, and tolerability of holybuvir and its metabolites and the effect of food on the PK of holybuvir and its metabolites in healthy Chinese subjects. A total of 96 subjects were enrolled in this study which included (i) a single-ascending-dose (SAD) study (100 to 1,200 mg), (ii) a food-effect (FE) study (600 mg), and (iii) a multiple-dose (MD) study (400 and 600 mg once daily for 14 days). The results showed that single oral administration of holybuvir at doses up to 1,200 mg was well tolerated. Holybuvir was rapidly absorbed and metabolized in the human body, which was consistent with the characteristics of holybuvir as a prodrug. PK analysis showed that Cmax and area under the curve (AUC) increased with dose in no dose-proportional manner after a single-dose administration (100 to 1,200 mg). Although high-fat meals did change the PK of holybuvir and its metabolites, clinical significance of changes in PK parameters induced by eating a high-fat diet would be further confirmed. Following multiple-dose administration, accumulation of metabolites SH229M4 and SH229M5-sul was observed. The favorable PK and safety results support the further development of holybuvir for patients with HCV. (This study was registered at Chinadrugtrials.org under identifier CTR20170859.).

Unexpectedly higher levels of anti-orthopoxvirus neutralizing antibodies are observed among gay men than general adult population.

BACKGROUND: The confirmed cases in the current outbreak of Monkeypox are predominantly identified in the networks of men who have sex with men (MSM). The preexisting antibodies may profoundly impact the transmission of monkeypox virus (MPXV), however the current-day prevalence of antibodies against MPXV among gay men is not well characterized. METHODS: A cohort of gay men (n = 326) and a cohort of the general adult population (n = 295) were enrolled in this study. Binding antibodies responses against MPXV/vaccinia and neutralizing antibody responses against vaccinia virus (Tiantan strain) were measured. The antibody responses of these two cohorts were then compared, as well as the responses of individuals born before and in/after 1981 (when the smallpox vaccination ceased in China). Finally, the correlation between the anti-MPXV antibody responses and the anti-vaccinia antibody responses, and the associations between preexisting anti-orthopoxvirus antibody responses and the diagnosed sexually transmitted infections (STIs) in the MSM cohort were analyzed separately. RESULTS: Our data showed that binding antibodies against MPXV H3, A29, A35, E8, B6, M1 proteins and vaccinia whole-virus lysate could be detected in individuals born both before and in/after 1981, of which the prevalence of anti-vaccinia binding antibodies was significantly higher among individuals born before 1981 in the general population cohort. Moreover, we unexpectedly found that the positive rates of binding antibody responses against MPXV H3, A29, A35, E8 and M1 proteins were significantly lower among individuals of the MSM cohort born in/after 1981, but the positive rates of anti-MPXV B6 and anti-vaccinia neutralizing antibody responses were significantly higher among these individuals compared to those of age-matched participants in the general population cohort. Additionally, we demonstrated that the positive and negative rates of anti-MPXV antibody responses were associated with the anti-vaccinia antibody responses among individuals born before 1981 in the general population cohort, but no significant association was observed among individuals born in/after 1981 in both cohorts. The positive rates of both the binding and the neutralizing antibody responses were comparable between individuals with and without diagnosed STIs in the MSM cohort. CONCLUSIONS: Anti-MPXV and anti-vaccinia antibodies could be readily detected in an MSM cohort and a general population cohort. And a higher level of anti-vaccinia neutralizing antibody responses was observed among individuals who did not get vaccinated against smallpox in the MSM cohort compared to age-matched individuals in the general population cohort.